This proposal is to request funding for a competing renewal of GM62648 to further develop tools and automation for x-ray crystallography of biological molecules. These tools have wide benefit to the structural biology community as they provide improved and optimized approaches to x-ray data collection and processing, as well as improvements and new methods for researchers to interact with the complex instrumentation at synchrotron crystallography beamlines. The synchrotron beamlines provide the majority of x-ray data for structural biology, and enable the pursuit of ever more challenging structural projects such as the ribosome, RNA polymerase complexes, and integral membrane proteins. The development of highly-automated technologies and methods is critical to the productivity of the scientific community and is necessary to increase the high- throughput capabilities for structural genomics and drug discovery projects. Over the past four years, we have designed, implemented, and tested an automated crystal mounting and alignment system, integrated it into the beamline control system in sector 5 at the Advanced Light Source, extensively tested and improved it in response to the tests and user feedback, started an effort with the group of Paul Adams (LBNL) to more fully automate data collection and processing, and begun the collaborative propagation of the system to other synchrotron beamlines. For this renewal, we propose to focus on the following goals: 1) Upgrade of the design of the automated sample- handling system - finalizing hardware of first generation system and improvements in the gripper assembly, and the design, implement, and testing second-generation system with cartesian geometry;2) Improve and more fully automate crystal recognition and alignment;3) Improve and more fully automate the sample-tracking subsystem;4) Develop a user-directed remote measurement and information management capability. These technologies are generalizable to the other sites and to a broad range of user needs.